In vitro antimicrobial efficacy of a silver alginate dressing on burn wound isolates.

OBJECTIVE: To test the antimicrobial effectiveness of a silver alginate dressing on opportunistic pathogens, namely meticillin-sensitive Staphylococcus aureus (MSSA) and meticillin-resistant Staphylococcus aureus (MRSA), Klebsiella spp., Enterococcus faecalis, Enterococcus faecium, Pseudomonas aeruginosa, Escherichia coli, Enterobacter sakazakii, Enterobacter cloacae, Serratia marcescens, Chryseobacterium indologenes, Proteus vulgaris and Acinetobacter baumannii. METHOD: In total, 40 microorganisms were isolated from patients attending three burn centres in the US and evaluated for their susceptibility to a silver alginate wound dressing, employing a corrected zone of inhibition assay, conducted on Mueller Hinton agar (MHA). RESULTS: The sizes of the corrected zones of inhibition varied between and within genera. For example, all Acinetobacter baumannii strains were found to be sensitive to ionic silver at pH 7, with a mean of 2.8mm, compared with 3.5mm at pH 5.5. The silver alginate dressing also demonstrated activity on all strains of Enterobacter and Escherichia coli, with susceptibility to the silver alginate dressing enhanced at pH 5.5. For Enterococcus spp. the average corrected zone of inhibition at pH 7 was 3.6mm, versus 4.9mm at pH 5.5. All strains of Pseudomonas aeruginosa were found to be sensitive to the silver alginate dressing. The average corrected zone of inhibition was 6.9mm at pH 7, compared with 8mm at pH 5.5. For MRSA and Staphylococcus aureus, it ranged from 4.5mm to 7.5mm at pH 7. When the pH was decreased to 5.5, the corrected zone of inhibition increased. CONCLUSION: This study demonstrates the activity of a silver alginate dressing on a wide range of burn isolates, including antibiotic-resistant bacteria, isolated from three different burn centres in the US. It also highlights the possible importance of pH and its potential effects on antimicrobial performance and microbial susceptibility. However, more extensive testing is required to substantiate this. CONFLICT OF INTEREST: SLP is employed by Advanced Medical Solutions Ltd.

A comparison of the antimicrobial efficacy of two silver-containing wound dressings on burn wound isolates.

OBJECTIVE: To evaluate and compare the efficacy of a silver alginate (SA) dressing and a silver carboxymethyl cellulose (SCMC) dressing on burn isolates grown within the quasi/non-biofilm state and the biofilm phenotypic states. METHOD: Antimicrobial activity was tested using 46 burn wound isolates with a corrected zone of inhibition (CZOI) assay on agar (quasi/non-biofilm) and poloxamer (biofilm). RESULTS: All Gram-negative and positive isolates evaluated were found to be sensitive to both silver containing wound dressings, although superior antimicrobial activity was observed for a select number of specific bacteria when grown in the quasi/non-biofilm phenotypic state, for the SCMC dressing. However, the majority of isolates demonstrated reduced sensitivity to silver when grown as a biofilm, compared with growth in the quasi/non-biofilm phenotypic state. Both dressings demonstrated equivalent antimicrobial activity on Gram-negative isolates grown in the biofilm phenotypic state. For the Gram-positive isolates growing in the biofilm phenotypic state, there appeared to be greater sensitivity to the SA dressing compared with the SCMC dressing, although this result was not statistically significant. CONCLUSION: This study demonstrated the antimicrobial effectiveness of an SA and SCMC dressing in inhibiting the growth of burn isolates residing in both the quasi/non-biofilm and biofilm phenotypic states. It also suggests the SA dressing could be more effective on Gram-positive isolates grown in the biofilm phenotypic state, compared with SCMC dressing.

Polymer Brushes Containing Sulfonated Sugar Repeat Units: Synthesis, Characterization and In Vitro Testing of Blood Coagulation Activation.

A new polymer brush chemistry containing sulfonated carbohydrate repeat units has been synthesized from silicon substrates using ATRP methods and characterized both in bulk and using surface analysis. The polymer brush was designed to act as a mimic for the naturally occurring sulfonated glycosaminoglycan, heparin, commonly used for modifying blood-contacting surfaces both in vitro and in vivo. Surface analysis showed conversion of brush saccharide precursor chemistry to the desired sulfonated polymer product. The sulfonated polymer brush surface was further analyzed using three conventional in vitro tests for blood compatibility -- plasma recalcification times, complement activation, and thrombin generation. The sulfonated polymer brush films on silicon oxide wafers exhibited better assay performance in these blood component assays than the unsulfonated sugar functionalized polymer brush in all tests performed.

The spirochete FlaA periplasmic flagellar sheath protein impacts flagellar helicity.

Spirochete periplasmic flagella (PFs), including those from Brachyspira (Serpulina), Spirochaeta, Treponema, and Leptospira spp., have a unique structure. In most spirochete species, the periplasmic flagellar filaments consist of a core of at least three proteins (FlaB1, FlaB2, and FlaB3) and a sheath protein (FlaA). Each of these proteins is encoded by a separate gene. Using Brachyspira hyodysenteriae as a model system for analyzing PF function by allelic exchange mutagenesis, we analyzed purified PFs from previously constructed flaA::cat, flaA::kan, and flaB1::kan mutants and newly constructed flaB2::cat and flaB3::cat mutants. We investigated whether any of these mutants had a loss of motility and altered PF structure. As formerly found with flaA::cat, flaA::kan, and flaB1::kan mutants, flaB2::cat and flaB3::cat mutants were still motile, but all were less motile than the wild-type strain, using a swarm-plate assay. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis indicated that each mutation resulted in the specific loss of the cognate gene product in the assembled purified PFs. Consistent with these results, Northern blot analysis indicated that each flagellar filament gene was monocistronic. In contrast to previous results that analyzed PFs attached to disrupted cells, purified PFs from a flaA::cat mutant were significantly thinner (19.6 nm) than those of the wild-type strain and flaB1::kan, flaB2::cat, and flaB3::cat mutants (24 to 25 nm). These results provide supportive genetic evidence that FlaA forms a sheath around the FlaB core. Using high-magnification dark-field microscopy, we also found that flaA::cat and flaA::kan mutants produced PFs with a smaller helix pitch and helix diameter compared to the wild-type strain and flaB mutants. These results indicate that the interaction of FlaA with the FlaB core impacts periplasmic flagellar helical morphology.

Borrelia burgdorferi periplasmic flagella have both skeletal and motility functions.

Bacterial shape usually is dictated by the peptidoglycan layer of the cell wall. In this paper, we show that the morphology of the Lyme disease spirochete Borrelia burgdorferi is the result of a complex interaction between the cell cylinder and the internal periplasmic flagella. B. burgdorferi has a bundle of 7-11 helically shaped periplasmic flagella attached at each end of the cell cylinder and has a flat-wave cell morphology. Backward moving, propagating waves enable these bacteria to swim in both low viscosity media and highly viscous gel-like media. Using targeted mutagenesis, we inactivated the gene encoding the major periplasmic flagellar filament protein FlaB. The resulting flaB mutants not only were nonmotile, but were rod-shaped. Western blot analysis indicated that FlaB was no longer synthesized, and electron microscopy revealed that the mutants were completely deficient in periplasmic flagella. Wild-type cells poisoned with the protonophore carbonyl cyanide-m-chlorophenylhydrazone retained their flat-wave morphology, indicating that the periplasmic flagella do not need to be energized for the cell to maintain this shape. Our results indicate that the periplasmic flagella of B. burgdorferi have a skeletal function. These organelles dynamically interact with the rod-shaped cell cylinder to enable the cell to swim, and to confer in part its flat-wave morphology.

Structure and expression of the FlaA periplasmic flagellar protein of Borrelia burgdorferi.

The spirochete which causes Lyme disease, Borrelia burgdorferi, has many features common to other spirochete species. Outermost is a membrane sheath, and within this sheath are the cell cylinder and periplasmic flagella (PFs). The PFs are subterminally attached to the cell cylinder and overlap in the center of the cell. Most descriptions of the B. burgdorferi flagellar filaments indicate that these organelles consist of only one flagellin protein (FlaB). In contrast, the PFs from other spirochete species are comprised of an outer layer of FlaA and a core of FlaB. We recently found that a flaA homolog was expressed in B. burgdorferi and that it mapped in a fla/che operon. These results led us to analyze the PFs and FlaA of B. burgdorferi in detail. Using Triton X-100 to remove the outer membrane and isolate the PFs, we found that the 38.0-kDa FlaA protein purified with the PFs in association with the 41.0-kDa FlaB protein. On the other hand, purifying the PFs by using Sarkosyl resulted in no FlaA in the isolated PFs. Sarkosyl has been used by others to purify B. burgdorferi PFs, and our results explain in part their failure to find FlaA. Unlike other spirochetes, B. burgdorferi FlaA was expressed at a lower level than FlaB. In characterizing FlaA, we found that it was posttranslationally modified by glycosylation, and thus it resembles its counterpart from Serpulina hyodysenteriae. We also tested if FlaA was synthesized in a spontaneously occurring PF mutant of B. burgdorferi (HB19Fla-). Although this mutant still synthesized flaA message in amounts similar to the wild-type amounts, it failed to synthesize FlaA protein. These results suggest that, in agreement with data found for FlaB and other spirochete flagellar proteins, FlaA is likely to be regulated on the translational level. Western blot analysis using Treponema pallidum anti-FlaA serum indicated that FlaA was antigenically well conserved in several spirochete species. Taken together, the results indicate that both FlaA and FlaB comprise the PFs of B. burgdorferi and that they are regulated differently from flagellin proteins of other bacteria.